α-Adrenergic Control of Volume-Regulated Cl − Currents in Rabbit Atrial Myocytes

Abstract

Abstract α-Adrenergic stimulation is known to play a role in cardiac arrhythmogenesis and to modulate a variety of cardiac K + currents. The effects of α-adrenergic stimulation on Cl − currents are largely unknown. Many cardiac cell types show a volume-sensitive Cl − current induced by cell swelling (I Cl.swell ). The present experiments were designed to assess the potential α-adrenergic modulation of I Cl.swell in rabbit atrial myocytes. I Cl.swell was induced with the use of a hypotonic superfusate, under conditions designed to prevent currents carried by K + , Na + , and Ca 2+ ions. A basal Cl − current (I Cl.b ) was observed under isotonic conditions in 128 of 150 cells (85%), had the same dependency on [Cl − ] o as I Cl.swell , and was reduced by cell shrinkage induced by hypertonic superfusion, suggesting that I Cl.b is carried by the same volume-sensitive Cl − conductance as I Cl.swell . Phenylephrine produced a concentration-dependent and near-complete inhibition of I Cl.b and I Cl.swell , with EC 50 values of 86±5 and 72±7 (mean±SEM) μmol/L, respectively, at +20 mV. Norepinephrine (administered in the presence of 1 μmol/L propranolol) also inhibited I Cl.b and I Cl.swell , with EC 50 values of 2.6±0.1 and 2.8±0.4 μmol/L, respectively. The concentration-response curve for phenylephrine was shifted significantly ( P <.001) to the right by the α 1 -adrenoceptor antagonist prazosin and by the α 1A -receptor antagonists (+)-niguldipine and 5-methylurapidil but was unaltered by the α 1B -receptor antagonist chloroethylclonidine (100 μmol/L). Inhibition of protein kinase C (PKC) with staurosporine, H-7, or 18-hour preincubation with the phorbol ester 4β-phorbol 12-myristate 13-acetate (PMA, 500 nmol/L) blocked the effects of phenylephrine on I Cl.swell , and the highly selective PKC inhibitor bisindolylmaleimide blocked the effects of norepinephrine on I Cl.swell and I Cl.b . Both PMA and 1-oleoyl-2-acetylglycerol inhibited I Cl.swell in a concentration-dependent fashion. In blinded studies, the phorbol ester phorbol 12,13-didecanoate (PDD) reduced I Cl.swell by 91±3%; its inactive analogue 4α-PDD had no effect (mean change, 3±1%). Preincubation with pertussis toxin (PTX) prevented the actions of phenylephrine on I Cl.swell , indicating a role for a PTX-sensitive guanine nucleotide–binding (G) protein. We conclude that α-adrenergic agonists inhibit volume-sensitive Cl − currents in rabbit atrial cells by interacting with an α 1A -adrenoceptor mechanism that is coupled to PKC via a PTX-sensitive G protein. These results suggest a potentially novel mechanism of α-adrenergic control of cardiac electrical activity, the inhibition of volume-sensitive Cl − currents, and indicate that PKC, well known to elicit phosphorylation-dependent Cl − currents in cat and guinea pig ventricular myocytes, is also capable of potently inhibiting other forms of cardiac Cl − current.